FasCure Therapeutics focuses on the development of adjuvant systems to generate prophylactic and therapeutic immune responses. The major objective of this Phase 1 STTR proposal is to reformulate a lead plague subunit vaccine with a novel adjuvant system to improve its protective efficacy. Yersinia pestis is the causative agent of bubonic and pneumonic plague and a potential bioweapon. Currently there is not a vaccine available to protect the public from a potential epidemic or bioterrorism event. While Y. pestis infection can be treated with antibiotics, the effective treatment window for primary pneumonic infection is very short (less than 24 hrs. after exposure). Furthermore, naturally acquired resistance to antibiotics has been reported, and weaponized Y. pestis could likely be modified to be resistant to antibiotic treatment. Therefore, the development of an efficacious plague vaccine is an important scientific endeavor and public policy goal. The current lead vaccine against plague is a subunit vaccine formulation consisting of a recombinant F1- LcrV fusion protein (rF1-V) adjuvanted with alum. This vaccine has proven effective in rodent models, but has decreased efficacy in primate models and variable antibody production in clinical trials. Therefore, while rF1-V appears to be an excellent vaccine candidate for plague, it needs further improvement to produce a better immunogenicity profile. While the current vaccine primarily generates a Th2 humoral immune response, several studies have demonstrated a critical role of Th1 cellular responses in protection against plague. Therefore, the primary focus of this proposal is to establish a novel adjuvant system that generate a mixed Th1 and Th2 immune responses, and use adjuvant to improve the efficacy of lead rF1V vaccine. This goal will be achieved by using alum, MPL, and SA-4-1BBL as agonists NLRs, TLR4, and the 4-1BB receptor, respectively, that are critical for the generation of effective Th1 cellular and Th2 humoral immune responses. Aim 1 will establish the optimum dose of each adjuvant to generate balanced, robust Th1 and Th2 responses against the rF1-V antigen. Aim 2 will use the lead vaccine formulation to determine its protective efficacy against pneumonic plague in mice. If successful, a Phase II proposal will be submitted to further develop this vaccine formulation by testing its protective efficacy against plague using a nonhuman primate model as a prelude to clinical trials. Importantly, the proposed adjuvant system may also be used as a platform to generate new subunit vaccines, or improve the efficacy of the existing ones, against other intracellular pathogens that need a balanced immune response.